Interpretive Summary: Foot-and-mouth disease virus (FMDV) causes an economically devastating disease of cloven-hoofed animals including swine and cattle. Eliminating FMD outbreaks in previously disease-free countries often relies on restriction of animal movement, massive slaughter of infected and in contact susceptible animals, and vaccination. The virus replicates very rapidly within the infected animal and spreads among in-contact susceptible animals. Disease signs can appear as early as 2 days post exposure to the virus. Thus, early protection is required in the event of an FMD outbreak in a disease-free country such as the US. However, the current inactivated whole virus vaccine and new subunit vaccines that are being developed cannot induce complete protection prior to 7 days postvaccination. To attempt to induce rapid protection, we explored the possibility of using RNA interference (RNAi) as a novel anti-FMD approach. RNAi is a natural process by which double-stranded RNA directs the inhibition of gene expression. Over the past couple of years several laboratories have used this technology to attenuate viral infection in cell culture. In this manuscript we demonstrate that RNAi inhibits the expression of FMDV RNA and protein in infected porcine cells. Furthermore, the amount of virus produced was reduced by approximately 97-98%. Since FMDV is a highly variable virus it is necessary to demonstrate that an antiviral strategy be effective against many different virus serotypes. In this study we demonstrated that RNAi reduced the amount of virus produced in cells infected with 4 different virus serotypes. These results highlight the potential of RNAi as a new virus control strategy.

Technical Abstract:
Foot-and-mouth disease virus (FMDV) is one of the most contagious agents of animals. Recent outbreaks in disease-free countries have prompted to the development of new control strategies that could improve the levels of protection against this virus. In an attempt to test whether FMDV could be targeted by small interfering RNAs (siRNAs) we have delivered to porcine cells short hairpin RNAs (shRNAs) directed against viral targets. Treatment of IBRS-2 cells with plasmids expressing shRNAs caused a significant reduction (>70%) in the synthesis of viral proteins and RNA, as well as a decrease in virus yield. The antiviral effect was sequence specific and not attributable to induction of the interferon pathway. We have identified a highly conserved sequence in the 2B nonstructural protein coding region as a prominent target for RNA interference. Infection of cells that expressed 2B shRNA with 3 FMDV serotypes resulted in efficient silencing.